Fig. 3

ALKBH5 abolishes migration/invasion capabilities of HCC cells in vitro and inhibits metastasis in vivo. a Transwell assays of Huh7, MHCC97H and HCCLM3 were applied to measure their migration and invasion abilities (scale bars, 200 μm). Bar charts showed the relative count (refer to negative control group) of cells which passed through the chamber membrane in each group (right); b Wound healing assays were conducted to compare the migration capabilities of three HCC cells after silencing or overexpression of ALKBH5. The difference in cell margin between 0 h and 72 h showed the moving track of cells; The percentage of healed area was quantified (right); c and d Alterations of cytoskeleton represented with immunofluorescent imaging were detected under the knockdown of ALKBH5 in Huh7 (c) and MHCC97H (d) cells. Phalloidin (red color) was applied for cytoskeleton staining, while DAPI (blue color) was used to mark the nuclei (scale bars, 30 μm). A divergent pattern of cytoskeleton with slenderer microtubules or microfilaments and more pseudopodia indicated a more flexible migrating style of cells; e, f and g HCCLM3 cells transfected with ALKBH5-overexpressing or control vector lentiviruses were injected into mice via tail vain to establish pulmonary metastasis models (n = 5). Representative in vivo images of mice were taken with quantification of luciferase activity in the lung region (e). Metastatic tumor foci in lungs were photographed and quantified (f), and their presence was further confirmed by HE staining (g) (scale bars, 100 μm)